LOCUS       sequence-416700-        6565 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of HA-tagged human PPM1H S124A/S211A.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6565)
  AUTHORS   Wondwossen M Yeshaw , Ayan Adhikari, Claire Y Chiang, Herschel S 
            Dhekne, Paulina S Wawro, Suzanne R Pfeffer
  TITLE     Localization of PPM1H phosphatase tunes Parkinson's disease-linked 
            LRRK2 kinase-mediated Rab GTPase phosphorylation and ciliogenesis
  JOURNAL   Proc Natl Acad Sci U S A. 2023 Oct 31;120(44)
REFERENCE   2  (bases 1 to 6565)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Proc Natl 
            Acad Sci U S A. 2023 Oct 31;120(44)"
COMMENT     Sequence Label: pCMV5D HA-PPM1H S124A/S211A
FEATURES             Location/Qualifiers
     source          1..6565
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        110..489
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        490..693
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     644..664
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             738..764
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             2325..2351
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     primer_bind     complement(2392..2409)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2398..2622
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(2755..2774)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    2906..2953
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     CDS             2961..3983
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     polyA_signal    4113..4234
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4150..4169)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4204..4223
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4283..4299)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4283..4299)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4296..4318)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4307..4323
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(4331..4337,4338..4355,4356..4361))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4376..4397
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4514..4531)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4685..5273)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4765..4784)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5444..6235,6236..6304))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTIRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     6067..6086
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6305..6409)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6484..6503)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 aaaatttaag ctacaacaag gcaaggcttg accgacaatt gcatgaagaa tctgcttagg
       61 gttaggcgtt ttgcgctgct tcgcgatgta cgggccagat atacgcgttg acattgatta
      121 ttgactagtt attaatagta atcaattacg gggtcattag ttcatagccc atatatggag
      181 ttccgcgtta cataacttac ggtaaatggc ccgcctggct gaccgcccaa cgacccccgc
      241 ccattgacgt caataatgac gtatgttccc atagtaacgc caatagggac tttccattga
      301 cgtcaatggg tggagtattt acggtaaact gcccacttgg cagtacatca agtgtatcat
      361 atgccaagtc cgccccctat tgacgtcaat gacggtaaat ggcccgcctg gcattatgcc
      421 cagtacatga ccttacggga ctttcctact tggcagtaca tctacgtatt agtcatcgct
      481 attaccatgg tgatgcggtt ttggcagtac accaatgggc gtggatagcg gtttgactca
      541 cggggatttc caagtctcca ccccattgac gtcaatggga gtttgttttg gcaccaaaat
      601 caacgggact ttccaaaatg tcgtaataac cccgccccgt tgacgcaaat gggcggtagg
      661 cgtgtacggt gggaggtcta tataagcaga gctcgtttag tgaaccgtca gaattggcca
      721 ccggatctgc cgccatgtac ccatacgatg tgccagatta cgccggatcc atgctcactc
      781 gagtgaaatc tgccgtggcc aatttcatgg gcggcatcat ggctggcagc tcaggctccg
      841 agcacggcgg cggcagctgc ggaggctcgg acctgcccct gcgtttcccc tacgggcggc
      901 cagagttcct ggggctgtct caggacgagg tggagtgcag cgccgaccac atcgcccgcc
      961 ccatcctcat cctcaaggag actcggcggc tgccctgggc cactggctac gcagaggtta
     1021 tcaatgccgg gaagagcaca cacaatgaag accaagccag ctgtgaggtg ctcactgtga
     1081 agaagaaggc aggggccgtg acctcaaccc caaacaggaa ctcatccaag agacggtccg
     1141 cccttcccaa tggggaaggg ctgcagctga aggagaactc ggaatccgag ggtgtttcct
     1201 gccactattg gtcgctgttt gacgggcacg cggggtccgg ggccgcggtg gtggcgtcac
     1261 gcctgctgca gcaccacatc acggagcagc tgcaggacat cgtggacatc ctgaagaact
     1321 ccgccgtcct gccccctacc tgcctggggg aggagcctga gaacacgccc gccaacagcc
     1381 ggactctgac ccgggcagcc gccctgcgcg gaggggtggg ggccccgggc tcccccagca
     1441 cgccccccac acgcttcttt accgagaaga agattcccca tgagtgcctg gtcatcggag
     1501 cgcttgaaag tgcattcaag gaaatggacc tacagataga acgagagagg agttcatata
     1561 atatatctgg tggctgcacg gccctcattg tgatttgcct tttggggaag ctgtatgttg
     1621 caaatgctgg ggatagcagg gccataatca tcagaaatgg agaaattatc cccatgtctt
     1681 cagaatttac ccccgagacg gagcgccagc gacttcagta cctggcattc atgcagcctc
     1741 acttgctggg aaatgagttc acacatttgg agtttccaag gagagtacag agaaaggagc
     1801 ttggaaagaa gatgctctac agggacttta atatgacagg ctgggcatac aaaaccattg
     1861 aggatgagga cttgaagttc ccccttatat atggagaagg caagaaggcc cgggtaatgg
     1921 caactattgg agtgaccagg ggacttgggg accatgacct gaaggtgcat gactccaaca
     1981 tctacattaa accattcctg tcttcagctc cagaggtaag aatctacgat ctttcaaaat
     2041 atgatcatgg atcagatgat gtgctgatct tggccactga tggactctgg gacgttttat
     2101 caaatgaaga agtagcagaa gcaatcactc agtttcttcc taactgtgat ccagatgatc
     2161 ctcacaggta cacactggca gctcaggacc tggtgatgcg tgcccggggt gtgctgaagg
     2221 acagaggatg gcggatatct aatgaccgac tgggctcagg agacgacatt tctgtatatg
     2281 tcattccttt aatacatgga aacaagctgt catgagcggc cgcttaccca tacgatgtgc
     2341 cagattacgc ctaagtcgag tctagagggc ccgtttaaac ccgctgatca gcctcgactg
     2401 tgccttctag ttgccagcca tctgttgttt gcccctcccc cgtgccttcc ttgaccctgg
     2461 aaggtgccac tcccactgtc ctttcctaat aaaatgagga aattgcatcg cattgtctga
     2521 gtaggtgtca ttctattctg gggggtgggg tggggcagga cagcaagggg gaggattggg
     2581 aagacaatag caggcatgct ggggatgcgg tgggctctat ggcttctgag gcggaaagaa
     2641 ccagctgggg ctctaggggg tatccccacg cgccctgtag cggcgcatta agcgcggcgg
     2701 gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag cgccctagcg cccgctcctt
     2761 tcgctttctt cccttccttt ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc
     2821 gggggctccc tttagggttc cgatttagtg ctttacggca cctcgacccc aaaaaacttg
     2881 attagggtga tggttcacgt acctagaagt tcctattccg aagttcctat tctctagaaa
     2941 gtataggaac ttccttggcc aaaaagcctg aactcaccgc gacgtctgtc gagaagtttc
     3001 tgatcgaaaa gttcgacagc gtctccgacc tgatgcagct ctcggagggc gaagaatctc
     3061 gtgctttcag cttcgatgta ggagggcgtg gatatgtcct gcgggtaaat agctgcgccg
     3121 atggtttcta caaagatcgt tatgtttatc ggcactttgc atcggccgcg ctcccgattc
     3181 cggaagtgct tgacattggg gagttcagcg agagcctgac ctattgcatc tcccgccgtg
     3241 cacagggtgt cacgttgcaa gacctgcctg aaaccgaact gcccgctgtt ctgcagccgg
     3301 tcgcggaggc catggatgcg atcgctgcgg ccgatcttag ccagacgagc gggttcggcc
     3361 cattcggacc gcaaggaatc ggtcaataca ctacatggcg tgatttcata tgcgcgattg
     3421 ctgatcccca tgtgtatcac tggcaaactg tgatggacga caccgtcagt gcgtccgtcg
     3481 cgcaggctct cgatgagctg atgctttggg ccgaggactg ccccgaagtc cggcacctcg
     3541 tgcacgcgga tttcggctcc aacaatgtcc tgacggacaa tggccgcata acagcggtca
     3601 ttgactggag cgaggcgatg ttcggggatt cccaatacga ggtcgccaac atcttcttct
     3661 ggaggccgtg gttggcttgt atggagcagc agacgcgcta cttcgagcgg aggcatccgg
     3721 agcttgcagg atcgccgcgg ctccgggcgt atatgctccg cattggtctt gaccaactct
     3781 atcagagctt ggttgacggc aatttcgatg atgcagcttg ggcgcagggt cgatgcgacg
     3841 caatcgtccg atccggagcc gggactgtcg ggcgtacaca aatcgcccgc agaagcgcgg
     3901 ccgtctggac cgatggctgt gtagaagtac tcgccgatag tggaaaccga cgccccagca
     3961 ctcgtccgag ggcaaaggaa tagcacgtac tacgagattt cgattccacc gccgccttct
     4021 atgaaaggtt gggcttcgga atcgttttcc gggacgccgg ctggatgatc ctccagcgcg
     4081 gggatctcat gctggagttc ttcgcccacc ccaacttgtt tattgcagct tataatggtt
     4141 acaaataaag caatagcatc acaaatttca caaataaagc atttttttca ctgcattcta
     4201 gttgtggttt gtccaaactc atcaatgtat cttatcatgt ctgtataccg tcgacctcta
     4261 gctagagctt ggcgtaatca tggtcatagc tgtttcctgt gtgaaattgt tatccgctca
     4321 caattccaca caacatacga gccggaagca taaagtgtaa agcctggggt gcctaatgag
     4381 tgagctaact cacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt
     4441 cgtgccagct gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc
     4501 gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg cggcgagcgg
     4561 tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa
     4621 agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg
     4681 cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga
     4741 ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg
     4801 tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg
     4861 gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc
     4921 gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg
     4981 gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca
     5041 ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt
     5101 ggcctaacta cggctacact agaagaacag tatttggtat ctgcgctctg ctgaagccag
     5161 ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg
     5221 gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc
     5281 ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt
     5341 tggtcatgag attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt
     5401 ttaaatcaat ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca
     5461 gtgaggcacc tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg
     5521 tcgtgtagat aactacgata cgggagggct taccatctgg ccccagtgct gcaatgatac
     5581 cgcgagaccc acgctcaccg gctccagatt tatcagcaat aaaccagcca gccggaaggg
     5641 ccgagcgcag aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc
     5701 gggaagctag agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccattgcta
     5761 caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac
     5821 gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc
     5881 ctccgatcgt tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac
     5941 tgcataattc tcttattgtc atgccatccg taagatgctt ttctgtgact ggtgagtact
     6001 caaccaagtc attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa
     6061 tacgggataa taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt
     6121 cttcggggcg aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca
     6181 ctcgtgcacc caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa
     6241 aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac
     6301 tcatactctt cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg
     6361 gatacatatt tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc
     6421 gaaaagtgcc acctgacgtc gacggatcgg gagatctccc gatcccctat ggtgcactct
     6481 cagtacaatc tgctctgatg ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt
     6541 ggaggtcgct gagtagtgcg cgagc
//
